Endocytosis of the tachykinin neuropeptide, neurokinin B, in astrocytes and its role in cellular copper uptake

The tachykinin neuropeptide, neurokinin B (NKB), belongs to a family of peptides having diverse roles in the brain. NKB, along with several other tachykinins, has been identified as a copper-binding peptide, however the physiological relevance of the binding is unclear. Previously, NKB was shown to limit the ability of copper to enter astrocytes and disrupt calcium homeostasis and it was thought that the peptide was sequestering the metal extracellularly. Here we use a fluorescein-labelled NKB peptide (F-NKB) to show that NKB is not retained extracellularly, but is endocytosed within 10–20min after addition to the cell media. The endocytosis is not inhibited when NKB is delivered as a copper-complex, [CuII(F-NKB)2]. Endocytosis of NKB can increase intracellular copper. Comparison to cells cultured in copper-free buffer indicated that apo-NKB can facilitate uptake of copper found in normal culture media. To achieve this NKB must compete with a variety of copper proteins, and we show that NKB can successfully compete with copper-binding peptides derived from the prion protein, itself associated with Cu(II) and Zn(II) metabolism. We suggest a mechanism of receptor mediated endocytosis to account for the observations. Neurokinin B (NKB) outcompetes the prion protein (PrPC) for copper and undergoes rapid endocytosis, likely mediated by the G protein-coupled neurokinin 3 receptor receptor. This process suggests a mechanism for copper uptake that avoids cytosolic exposure and highlights how NKB could be involved in synaptic copper homeostasis. [Display omitted] •The neurokinin, NKB, is rapidly endocytosed in astrocyte cells.•Copper-coordination of NKB, forming [CuII(NKB)2], does not inhibit endocytosis.•NKB can increase the cellular copper concentration.•NKB preferentially binds copper in the presence of copper-binding prion peptides.